Target (protein/gene name): N-acylmannosamine kinaseNCBI Gene # or RefSeq#: Sequence ID: WP_000222983.1Protein ID (NP or XP #) or Wolbachia #: LN847353Organism (including strain): Streptococcus pneumoniae MLV-016Etiologic Risk Group (see link below):Disease Information:Pneumococcal disease is an infection caused by Streptococcus pneumoniae bacteria. These bacteria can cause many types of illnesses, including: pneumonia, ear infections, sinus infections, meningitis, and bacteremia. Pneumococcus bacteria are spread through coughing, sneezing, and close contact with an infected person. Symptoms of pneumococcal disease depend on the part of the body that is infected. They can include fever, cough, shortness of breath, chest pain, stiff neck, confusion and disorientation, sensitivity to light, joint pain, chills, ear pain, sleeplessness, and irritability. Pneumococcal disease occurs around the world. Outbreaks of pneumococcal disease are uncommon in countries that have introduced the pneumococcal conjugate vaccine, but may occur in certain situations, such as in nursing homes, childcare centers, or other institutions. Certain people are more likely to become ill with pneumococcal disease. This high-risk group includes adults 65 years of age or older and children younger than 2 years of age. People who have conditions that weaken the immune system, like diabetes, heart disease, lung disease, and HIV/AIDS, or people who smoke cigarettes or have asthma are also at increased risk for getting pneumococcal disease.Link to TDR Targets page: N/ALink to Gene Database page (NCBI, EuPath databases -e.g. TryTryp, PlasmoDB, etc - or PATRIC, etc.): https://blast.ncbi.nlm.nih.gov/Blast.cgi#alnHdr_882952178 Essentiality of this protein: Sialic acid (N-acetylneuraminic acid; NeuNAc) is one of the most important carbohydrates for Streptococcus pneumoniae due of its role as a carbon and energy source, receptor for adhesion and invasion and molecular signal for promotion of biofilm formation, nasopharyngeal carriage, and invasion of the lung. Is it a monomer or multimer as biological unit? (make prediction at http://www.ebi.ac.uk/msd-srv/prot_int/pistart.html): Monomer Complex of proteins?: N/A Druggable Target (list number or cite evidence from a paper/database showing druggable in another organism): “Many biological processes are controlled by cell surface sialylation. To gather further insight into these processes, methods to alter cell surface sialylation are required. One way to achieve this is inhibiting the key enzyme of silica acid de novo biosynthesis, the intracellular bifunctional UDP-N-acetylglucosamine epimerase/N-acetylmannosamine kinase (GNE/MNK).” http://www.nrcresearchpress.com/doi/abs/10.1139/cjc-2015-0603#.WQ_KmlLMy9Y EC#: 2.7.1.60Link to BRENDA EC# page: http://www.brenda-enzymes.org/enzyme.php?ecno=2.7.1.60Show screenshot of BRENDA enzyme mechanism schematic:Enzyme Assay information (spectrophotometric, coupled assay ?, reagents): Link to Sigma (or other company) page for assay (see Sigma links below): Or link (or citation) to paper that contains assay information: Links to assay reagents (substrates) pages: List cost and quantity of substrate reagents, supplier, and catalog #: Structure (PDB or Homology model):
PDB # or closest PDB entry if using homology model: 2YI1 For Homology Model option: Show pairwise alignment of your BLASTP search in NCBI against the PDB: Query Coverage: Max % Identities: % Positives: Chain used for homology: Current Inhibitors: Expression Information (has it been expressed in bacterial cells): Purification Method: Image of protein (PyMol with features delineated and shown separately): Amino Acid Sequence:MTYYVAIDIGGTNIKYGLVDQEGQLLESHEMPTEAHKGGPHILQKTKDIVASYLEKGPVAGVAISSAGMVDPDKGEIFYAGPQIPNYAGTQFKKEIEESFTIPCEIENDVNCAGLAEAVSGSGKGASVTLCLTIGTGIGGCLIMDRKVFHGFSNSACEVGYMHMQDGAFQDLASTTALVKYVAEAHGEDVDQWNGRRIFKEATEGNKICMEGIDRMVDYLGKGLANICYVANPEVVILGGGIMGQEAILKPKIRTALKEALVPSLAEKTRLEFAHHQNTAGMLGAYYHFKTKQS Length of your protein in Amino Acids: 294 Molecular Weight of your protein in kiloDaltons using the Expasy ProtParam website:Molar Extinction coefficient of your protein at 280 nm wavelength:TMpred graph Image (http://www.ch.embnet.org/software/TMPRED_form.html). Input your amino acid sequence to it. CDS Gene Sequence: ATGACATACTACGTTGCAATTGATATCGGTGGAACTAACATCAAGTATGGTTTGGTTGATCAAGAGGGGCAACTTCTTGAATCACATGAAATGCCAACAGAGGCGCATAAGGGTGGACCTCATATCTTACAAAAGACCAAAGATATCGTAGCTAGTTATTTAGAAAAAGGCCCAGTAGCAGGTGTTGCCATATCTTCTGCTGGGATGGTGGATCCGGATAAGGGTGAGATTTTCTATGCTGGGCCGCAAATCCCTAACTACGCAGGCACCCAGTTCAAAAAGGAAATCGAAGAAAGCTTTACTATTCCTTGTGAGATTGAAAATGATGTCAACTGTGCAGGTCTTGCTGAGGCAGTATCTGGTTCAGGCAAGGGAGCAAGTGTGACACTTTGCTTGACCATTGGAACCGGTATCGGTGGTTGCTTGATTATGGATAGGAAAGTCTTCCATGGTTTTAGTAATTCAGCCTGTGAAGTCGGGTATATGCATATGCAGGATGGAGCTTTTCAGGATCTCGCTTCTACAACAGCTTTAGTGAAATATGTAGCTGAAGCCCATGGAGAAGATGTTGATCAGTGGAATGGCCGACGTATCTTTAAGGAAGCTACTGAAGGAAACAAAATCTGCATGGAAGGTATTGACCGTATGGTTGACTATCTAGGAAAAGGTCTGGCAAATATTTGCTACGTTGCCAATCCAGAAGTGGTCATTCTTGGTGGTGGTATCATGGGGCAAGAGGCTATCCTCAAACCTAAGATCCGCACAGCCTTGAAAGAGGCTTTGGTACCAAGTTTAGCAGAAAAAACACGATTAGAATTTGCCCATCACCAAAATACAGCAGGGATGTTGGGTGCATATTATCATTTTAAGACAAAACAATCCTAG GC% Content for gene: CDS Gene Sequence (codon optimized) - copy from output of Primer Design Protocol: GC% Content for gene (codon optimized):
Do Not Need this info for Spring (but still copy these lines to your Target page for now)Primer design results for pNIC-Bsa4 cloning (list seqeunces of all of your ~40 nt long primers):(link to DNA Works output text file - that should be saved in your Google Docs folder after you did the primer design protocol)Ask a mentor, Dr. B, or a fellow researcher -how to link a GDocs file if you are not sure how to. Primer design results for 'tail' primers (this is just 2 sequences):
https://blast.ncbi.nlm.nih.gov/Blast.cgi#alnHdr_882952178
Essentiality of this protein: Sialic acid (N-acetylneuraminic acid; NeuNAc) is one of the most important carbohydrates for Streptococcus pneumoniae due of its role as a carbon and energy source, receptor for adhesion and invasion and molecular signal for promotion of biofilm formation, nasopharyngeal carriage, and invasion of the lung.
Is it a monomer or multimer as biological unit? (make prediction at http://www.ebi.ac.uk/msd-srv/prot_int/pistart.html): Monomer
Complex of proteins?: N/A
Druggable Target (list number or cite evidence from a paper/database showing druggable in another organism): “Many biological processes are controlled by cell surface sialylation. To gather further insight into these processes, methods to alter cell surface sialylation are required. One way to achieve this is inhibiting the key enzyme of silica acid de novo biosynthesis, the intracellular bifunctional UDP-N-acetylglucosamine epimerase/N-acetylmannosamine kinase (GNE/MNK).”
http://www.nrcresearchpress.com/doi/abs/10.1139/cjc-2015-0603#.WQ_KmlLMy9Y
EC#: 2.7.1.60Link to BRENDA EC# page: http://www.brenda-enzymes.org/enzyme.php?ecno=2.7.1.60Show screenshot of BRENDA enzyme mechanism schematic:Enzyme Assay information (spectrophotometric, coupled assay ?, reagents):
Link to Sigma (or other company) page for assay (see Sigma links below):
Or link (or citation) to paper that contains assay information:
Links to assay reagents (substrates) pages:
List cost and quantity of substrate reagents, supplier, and catalog #:
Structure (PDB or Homology model):
PDB # or closest PDB entry if using homology model: 2YI1
For Homology Model option:
Show pairwise alignment of your BLASTP search in NCBI against the PDB:
Query Coverage:
Max % Identities:
% Positives:
Chain used for homology:
Current Inhibitors:
Expression Information (has it been expressed in bacterial cells):
Purification Method:
Image of protein (PyMol with features delineated and shown separately):
Amino Acid Sequence: MTYYVAIDIGGTNIKYGLVDQEGQLLESHEMPTEAHKGGPHILQKTKDIVASYLEKGPVAGVAISSAGMVDPDKGEIFYAGPQIPNYAGTQFKKEIEESFTIPCEIENDVNCAGLAEAVSGSGKGASVTLCLTIGTGIGGCLIMDRKVFHGFSNSACEVGYMHMQDGAFQDLASTTALVKYVAEAHGEDVDQWNGRRIFKEATEGNKICMEGIDRMVDYLGKGLANICYVANPEVVILGGGIMGQEAILKPKIRTALKEALVPSLAEKTRLEFAHHQNTAGMLGAYYHFKTKQS
Length of your protein in Amino Acids: 294
Molecular Weight of your protein in kiloDaltons using the Expasy ProtParam website:Molar Extinction coefficient of your protein at 280 nm wavelength:TMpred graph Image (http://www.ch.embnet.org/software/TMPRED_form.html). Input your amino acid sequence to it.
CDS Gene Sequence:
ATGACATACTACGTTGCAATTGATATCGGTGGAACTAACATCAAGTATGGTTTGGTTGATCAAGAGGGGCAACTTCTTGAATCACATGAAATGCCAACAGAGGCGCATAAGGGTGGACCTCATATCTTACAAAAGACCAAAGATATCGTAGCTAGTTATTTAGAAAAAGGCCCAGTAGCAGGTGTTGCCATATCTTCTGCTGGGATGGTGGATCCGGATAAGGGTGAGATTTTCTATGCTGGGCCGCAAATCCCTAACTACGCAGGCACCCAGTTCAAAAAGGAAATCGAAGAAAGCTTTACTATTCCTTGTGAGATTGAAAATGATGTCAACTGTGCAGGTCTTGCTGAGGCAGTATCTGGTTCAGGCAAGGGAGCAAGTGTGACACTTTGCTTGACCATTGGAACCGGTATCGGTGGTTGCTTGATTATGGATAGGAAAGTCTTCCATGGTTTTAGTAATTCAGCCTGTGAAGTCGGGTATATGCATATGCAGGATGGAGCTTTTCAGGATCTCGCTTCTACAACAGCTTTAGTGAAATATGTAGCTGAAGCCCATGGAGAAGATGTTGATCAGTGGAATGGCCGACGTATCTTTAAGGAAGCTACTGAAGGAAACAAAATCTGCATGGAAGGTATTGACCGTATGGTTGACTATCTAGGAAAAGGTCTGGCAAATATTTGCTACGTTGCCAATCCAGAAGTGGTCATTCTTGGTGGTGGTATCATGGGGCAAGAGGCTATCCTCAAACCTAAGATCCGCACAGCCTTGAAAGAGGCTTTGGTACCAAGTTTAGCAGAAAAAACACGATTAGAATTTGCCCATCACCAAAATACAGCAGGGATGTTGGGTGCATATTATCATTTTAAGACAAAACAATCCTAG
GC% Content for gene:
CDS Gene Sequence (codon optimized) - copy from output of Primer Design Protocol:
GC% Content for gene (codon optimized):
Do Not Need this info for Spring (but still copy these lines to your Target page for now)Primer design results for pNIC-Bsa4 cloning (list seqeunces of all of your ~40 nt long primers):(link to DNA Works output text file - that should be saved in your Google Docs folder after you did the primer design protocol)Ask a mentor, Dr. B, or a fellow researcher -how to link a GDocs file if you are not sure how to.
Primer design results for 'tail' primers (this is just 2 sequences):